Rubberlike
protein hydrogels are unique in their remarkable stretchability
and resilience but are usually low in strength due to the largely
unstructured nature of the constitutive protein chains, which limits
their applications. Thus, reinforcing protein hydrogels while retaining
their rubberlike properties is of great interest and has remained
difficult to achieve. Here, we propose a fibrillization strategy to
reinforce hydrogels from engineered protein copolymers with photo-cross-linkable
resilin-like blocks and fibrillizable silklike blocks. First, the
designer copolymers with an increased ratio of the silk to resilin
blocks were photochemically cross-linked into rubberlike hydrogels
with reinforced mechanical properties. The increased silk-to-resilin
ratio also enabled self-assembly of the resulting copolymers into
fibrils in a time-dependent manner. This allowed controllable fibrillization
of the copolymer solutions at the supramolecular level for subsequent
photo-cross-linking into reinforced hydrogels. Alternatively, the
as-prepared chemically cross-linked hydrogels could be reinforced
at the material level by inducing fibrillization of the constitutive
protein chains. Finally, we demonstrated the advantage of reinforcing
these hydrogels for use as piezoresistive sensors to achieve an expanded
pressure detection range. We anticipate that this strategy may provide
intriguing opportunities to generate robust rubberlike biomaterials
for broad applications.
A strategy for photoinduced covalent immobilization of proteins on phenol-functionalized surfaces is described. Under visible light irradiation, the reaction can be completed within seconds at ambient temperature, with high yields in aqueous solution of physiological conditions. Protein immobilization is based on a ruthenium-catalyzed radical cross-linking reaction between proteins and phenol-modified surfaces, and the process has proven mild enough for lipase, Staphylococcus aureus protein A, and streptavidin to preserve their bioactivity. This strategy was successfully applied to antibody immobilization on different material platforms, including agarose beads, cellulose membranes, and glass wafers, thus providing a generic procedure for rapid biomodification of surfaces.
Matrix
stiffness and fibrous structure provided by the native extracellular
matrix have been increasingly appreciated as important cues in regulating
cell behaviors. Recapitulating these physical cues for cell fate regulation
remains a challenge due to the inherent difficulties in making mimetic
hydrogels with well-defined compositions, tunable stiffness, and structures.
Here, we present two series of fibrous and porous hydrogels with tunable
stiffness based on genetically engineered resilin-silk-like and resilin-like
protein polymers. Using these hydrogels as substrates, the mechanoresponses
of bone marrow mesenchymal stem cells to stiffness and fibrous structure
were systematically studied. For both hydrogel series, increasing
compression modulus from 8.5 to 14.5 and 23 kPa consistently promoted
cell proliferation and differentiation. Nonetheless, the promoting
effects were more pronounced on the fibrous gels than their porous
counterparts at all three stiffness levels. More interestingly, even
the softest fibrous gel (8.5 kPa) allowed the stem cells to exhibit
higher endothelial differentiation capability than the toughest porous
gel (23 kPa). The predominant role of fibrous structure on the synergistic
regulation of endothelial differentiation was further explored. It
was found that the stiffness signal activated Yes-associated protein
(YAP), the main regulator of endothelial differentiation, via spreading
of focal adhesions, whereas fibrous structure reinforced YAP activation
by promoting the maturation of focal adhesions and associated F-actin
alignment. Therefore, our results shed light on the interplay of physical
cues in regulating stem cells and may guide the fabrication of designer
proteinaceous matrices toward regenerative medicine.
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